This invention relates generally to the isolation of high-activity heparin and, more particularly, to the separation of heparin into fractions of relatively high specific activity and relatively low specific activity by affinity chromatography.
Heparin is a highly sulphated dextrorotatory mucopolysaccharide which is commonly employed in various biomedical applications for its anticoagulant properties in prolonging the clotting time of whole blood. It is typically isolated from mammalian tissue, such as animal livers, lungs or intestines, and thereafter purified by methods well known in the art. The anticoagulant potency of the isolated and purified heparin, is generally referred to as its "specific activity" and expressed as IU/mg.
Commercially available heparin typically has a specific activity of approximately 150 IU/mg, and has previously been shown to be composed of a mixture of different heparin species varying in their specific activities. Recent use of heparin as a coating for biomaterials exposed to blood has prompted a search for methods capable of separating the most active fractions. Several different techniques have previously been proposed for effecting a separation of the heparin of relatively high specific activity out of its mixture with heparin of relatively low specific activity, including gel filtration, sucrose density gradient centrifugation after binding with antithrombin III, affinity chromatography on antithrombin III-agarose, ion-exchange chromatography on DEAE-cellulose, and electrofocusing. However, all of these procedures permit fractionation of only small quantities of heparin, and have not proved to be practical for the large scale isolation of high-activity heparin in high yield.